Elsevier

Environmental Pollution

Volume 224, May 2017, Pages 706-713
Environmental Pollution

Hepatotoxicity of benzotriazole and its effect on the cadmium induced toxicity in zebrafish Danio rerio

https://doi.org/10.1016/j.envpol.2017.02.055Get rights and content

Highlights

  • 5.0μM of 1H-BTR induced no acute physiology toxic effects on zebrafish embryo.

  • 1H-BTR induced increasing expression of liver-specific fatty acid binding protein.

  • Co-exposure to 1H-BTR could decrease the toxicity of Cd in zebrafish.

  • Cd bioaccumulation increased significantly in zebrafish pre-exposed to 1H-BTR.

Abstract

As an emerging contaminant, 1-H-benzotriazole (1H-BTR) has been detected in the engineered and natural aquatic environments, which usually coexists with heavy metals and causes combined pollution. In the present study, wild-type and transgenic zebrafish Danio rerio were used to explore the acute toxicity as well as the single and joint hepatotoxicity of cadmium (Cd) and 1H-BTR. Although the acute toxicity of 1H-BTR to zebrafish was low, increased expression of liver-specific fatty acid binding protein was observed in transgenic zebrafish when the embryos were exposed to 5.0 μM of 1H-BTR for 30 days. Besides, co-exposure to 1H-BTR not only reduced the acute toxic effects induced by Cd, but also alleviated the Cd-induced liver atrophy in transgenic fish. Correspondingly, effects of combined exposure to 1H-BTR on the Cd-induced expressions of several signal pathway-related genes and superoxide dismutase and glutathione-s-transferase proteins were studied. Based on the determination of Cd bioaccumulation in fish and the complexing stability constant (β) of Cd-BTR complex in solution, the detoxification mechanism of co-existing 1H-BTR on Cd to the zebrafish was discussed.

Introduction

1-H-benzotriazole (1H-BTR) is widely used as corrosion inhibitor in engine antifreeze and printing inks (Avagyan et al., 2015, Liang et al., 2014). Reportedly, at least 9000 tons of BTR compounds were produced in the USA in 1999 (Hart et al., 2004). Given the high solubility of 1H-BTR in water (Table S1 in the Supplementary data), they can directly enter various aquatic ecosystems through municipal wastewater effluents and surface runoffs. Widespread occurrence of 1H-BTR in freshwater and marine environments has been reported (Seeland et al., 2012, Wang et al., 2016), occasionally at high levels of μg/L or even mg/L (Cancilla et al., 1998, Giger et al., 2006).

The reported sub-lethal impacts and sub-inhibitory effects of 1H-BTR to aquatic organisms and animals are commonly low. As found in the acute tests with Daphnia magna (Seeland et al., 2012), the 48 h EC10 and EC50 values of 1H-BTR were 3.94 and 107 mg/L, while in the 21-day reproduction tests 1H-BTR showed no adverse effect. Kadar et al. (2010) reported that the lowest observed effective concentration of 1H-BTR for the 48 h hatch rate in Ciona intestinalis was 32 mg/L (Kadar et al., 2010). However, more knowledge about 1H-BTR's physiological toxicity, such as hepatotoxicity to the aquatic organisms, is still needed (Liang et al., 2014).

Combined pollutants usually coexist in the engineered and natural aquatic environments, which might result in complicated toxic effect. For example, cadmium (Cd)-phenanthrene mixtures produced a less-than additive mortality in the oligochaete, Ilyodrilus templetoni in sediments (Gust and Fleeger, 2006), while a more-than-additive lethality induced by Cd-phenanthrene co-exposure was found in the waterborne copepod, Amphiascoides atopus (Fleeger et al., 2007). 1H-BTR is applied to retard the corrosion of metal surfaces by forming metal-BTR complexes (Health Council of the Netherlands, 2000). Although both of 1H-BTR and heavy metals are frequently detected in the aquatic environments, little is known about their combined toxic effects on the aquatic organisms.

In this study, wild-type and transgenic zebrafish (Danio rerio) were chosen as the test organisms, and the acute and developmental toxicity of 1H-BTR and cadmium (Cd) in their single and combined exposure treatments were assessed. The expressions of the genes related several typical pathways and the oxidative stress-related enzymes in liver of zebrafish in different exposure treatments were contrasted. Based on the measured distribution of Cd species in 1H-BTR solution, joint toxicity of co-existing Cd and 1H-BTR was further explained.

Section snippets

Chemicals

1H-BTR and cadmium chloride (CdCl2) were purchased from Sigma-Aldrich (>99.9% purity) and dissolved in egg water (60 mg/L sea salt dissolved in reconstituted water). The reconstituted purified water was prepared according to the ISO standard (7346-3: 1996), with pH of 8.0 and the hardness of approximately 150 mg/L of calcium carbonate (CaCO3).

Zebrafish

Two kinds of zebrafish (Danio rerio) were applied in toxic test of this study. Wild-type zebrafish were purchased from local ornamental fish suppliers.

Toxicity of Cd and 1H-BTR on the wild-type zebrafish embryo

According to the pre-experiment, 24 hpf blood-circulation inhibition, 56 hpf hatch rate inhibition, and 56 hpf survival rate of the wild-type zebrafish embryo were selected as the observed endpoints in this experiment. At each of the three endpoints, significant toxic effects of Cd (1.0 μM, single exposure) were found (p < 0.05) (Fig. 1). However, no significant effect at any endpoint was found when the embryos were exposed to 5.0 μM of 1H-BTR. Notably, when 1H-BTR (5.0 μM) presented

Low-dose effects of 1H-BTR

Due to the widespread application, BTR and derivatives were frequently detected in aquatic environments (Seeland et al., 2012), sometimes at very high concentrations. For example, concentration of methyl-BTR, up to over 2000 μg/L, was reported in the surface waters which related to airport runoffs (Corsi et al., 2003). In embryo of wild-type zebrafish, no developmental toxicity effect was observed with 4-day exposure of 1H-BTR at 5.0 μM (600 μg/L approximately). Liver plays an important role in

Conclusions

Although the acute toxicity of 1H-BTR was verified to be low, its hepatotoxicity, including changes of liver size and abnormal expression of typical genes and enzymes in liver was observed in zebrafish exposed to 1H-BTR at environmental concentration level. In addition, detoxification of Cd in zebrafish by the co-existing 1H-BTR was found, while this detoxification effect was not consistent with the changes of Cd bioaccumulation in fish. Based on the result of Cd species analysis, the

Acknowledgements

This work was supported by the Tianjin Talent Development Scholarship Fund in China (Grant No. [2013]52); the Tianjin Research Program of Application Foundation and Advanced Technology (14JCYBJC23200); and the Natural Science Foundation of China (Grant No. 21177065). We also thank Dr Xueyuan Gu of Nanjing University for her help in analyzing the potentiometric titration data. The authors declare that there are no conflicts of interest, and any studies involving fish and embryos were conducted

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